Molecular characterization and incidence of new tospovirus: Soybean Vein Necrosis Virus (SVNV) in Egypt / Caracterização molecular e incidência de novo tospovírus: Vírus da Necrose da Veia da Soja (SVNV) no Egito

Abstract Field survey study was conducted season (2017). Soybeans and weeds were weekly sampled randomly. Thrips adults were identified and counted. Detection of the virus isolate and the natural incidence was determined using; Mechanical transmission, host range, DAS-ELISA, RT-PCR. The natural incidence thrips individuals was detected depending on the SVNV% in thrips individuals and weeds hosts. Ten thrips species were associated with soybean plants in the field. The most abundant species was T. tabaci, average 256.5 average no.of individuals, followed by F. occidentalis (142.5 average no. of individuals), then N. variabilis (86.6/ average no. of individuals). Fourteen thrips species occurred on 5 legumes field crops and 41 weed plant species within soybean field. The highest average number 40.6.of individuals were recorded on Ammi majus. While the lowest one 3.3 average no. of individuals were on Urtica urens. Only 21diagnostic plant species were susceptible to infection with SVNV. G. max and Vigna radiate, were the highest percentage of infection 80% followed by V. unguilata & N. benthamiana, 75%. Egyptian isolate of Soybean vein necrosis virus (SVNV) in this study showed a high degree of similarity and it is closely related to TSWV from Egypt (DQ479968) and TCSV from USA (KY820965) with nucleotide sequence identity of 78%. Four thrips species transmitted SVNV (F. fusca 4.0%, F. schultzei 4.3%, F. tritici 3.3% and N. variabilis 68.0% transmission). Both C. phaseoli and M. sjostedti can acquire the virus but unable to transmit it. The following species; T. tabaci, F. occidentalis, S. dorsallis and T. palmi cannot acquire or transmit SVNV. The incidence of SVNV in the field started by the end of July then increased gradualy from 12.7 to 71.3% by the end of the season. In conclusion, few thrips individuals invaded soybean crops are enough to transmit high rate of SVNV within the crop. Furthermore, several vector species are also abundant on weeds, which are the major sources of soybean viruses transmitted to the crops. This information might be important for control and reduce the incidence of SVNV infection.

Resumo O estudo de pesquisa de campo foi realizado na temporada (2017). A soja e as ervas daninhas foram amostradas semanalmente de forma aleatória. Tripes adultos foram identificados e contados. A detecção do vírus isolado e a incidência natural foram determinadas usando transmissão mecânica, gama de hospedeiros, DAS-ELISA, RT-PCR. A incidência natural de tripes em indivíduos foi detectada dependendo da % de SVNV em tripes e hospedeiros infestantes. Dez espécies de tripes foram associadas a plantas de soja no campo. A espécie mais abundante foi T. tabaci, com média de 256,5 número médio de indivíduos, seguida por F. occidentalis (142,5) e N. variabilis (86,6 / número médio de indivíduos). Catorze espécies de tripes ocorreram em 5 culturas de leguminosas e 41 espécies de plantas daninhas dentro de campos de soja. O maior número médio de 40,6 indivíduos foi registrado em Ammi majus. Enquanto o mais baixo, 3,3 número médio de indivíduos, foi no Urtica urens. Apenas 21 espécies de plantas diagnosticadas foram suscetíveis à infecção com SVNV. G. max e Vigna radiate foram os maiores percentuais de infecção, 80%, seguidos por V. unguilata e N. benthamiana, 75%. O isolado egípcio neste estudo mostrou um alto grau de similaridade e está intimamente relacionado ao TSWV do Egito (DQ479968) e ao TCSV dos EUA (KY820965), com identidade de sequência de nucleotídeos de 78%. Quatro espécies de tripes transmitiram SVNV (F. fusca 4,0%, F. schultzei 4,3%, F. tritici 3,3% e N. variabilis 68,0% de transmissão). Tanto C. phaseoli quanto M. sjostedti podem adquirir o vírus, mas não podem transmiti-lo. As seguintes espécies, T. tabaci, F. occidentalis, S. dorsallis e T. palmi não podem adquirir ou transmitir SVNV. A incidência de SVNV no campo, iniciada no final de julho, aumentou gradativamente de 12,7 para 71,3% no final da temporada. Em conclusão, poucos indivíduos de tripes invadiram a cultura da soja e são suficientes para transmitir alta taxa de SVNV dentro da cultura. Além disso, várias espécies de vetores também abundam em ervas daninhas, que são as principais fontes dos vírus da soja transmitidos às lavouras. Essas informações podem ser importantes para controlar e reduzir a incidência de infecção por SVNV.

Elimination of TSWV from impatiens hawkerii Bull. and regeneration of virus-free plant

The possibility for obtaining virus free plants from Impatiens hawkerii Bull. shoots infected with Tomato spotted wilt virus (TSWV) through meristem-tip culture was examined. TSWV presence in I. hawkerii plants was detected by DAS-ELISA and RT-PCR and identification of the virus was confirmed by sequencing one of the chosen isolate (GenBank Accesion CQ132190). Meristem-tip explants (0.3-1.5 mm) from virus-infected shoots are cultured on MS media supplemented with different concentrations of the cytokinins, CPPU or TDZ (0.01-1.0 uM), respectively. Using this system, a large number of in vitro shoots could be produced from a single explant. Also, cytokinins showed a stimulatory effect on the length, fresh and dry weights of the newly formed shoots. Plant pigments content in I. hawkerii shoots increased significantly in the presence of cytokinins. Rooting of shoots was spontaneous on the same media. Rooted plantlets were transferred to soil where 97 percent successfully acclimatized. By DAS-ELISA and RT-PCR, 80 percent of the in vitro plantlets were shown to be a virus-free. Considering these, the present protocol seems to be an efficient method for in vitro generation of virus-free I. hawkerii plantlets by meristem tip cultures.

Correlations between symptoms and DAS - ELISA values in two sources of resistance against tomato spotted wilt virus

The commercial tomato, "Lycopersicon esculentum", cultivars Ângela Gigante I-5100 and Santa Clara (tsopovirus susceptible), the Stevens cultivar, lines and experimental hybrids (tospovirus resistant), were used : a) to evaluate two sources of resistance to tospoviruses in tomato (one from "L. esculentum"/Rey de Los Tempranos, and another one from "L. peruvianum"/Stevens); and b) to verify statistically significant correlation between virus concentration determined through DAS-ELISA and symptomatology. In the Ângela GiganteI-5100 and the Santa Clara cultivars, the correlation coefficient was highly significant and the symptom average level was high, showing that in susceptible cultivars the symptom evolution is related to the virus multiplication in the plant. As for the TOM 547 and TOM 556 lines (background of Ângela Gigante I-5100 and Santa Clara, respectively), there was no statistical correlation between the symptoms and the viral particule concentration. Plants with light virus symptoms showed high absorbance values. Consequently we may assume that the resistance with 'Rey de Los Tempranos' background may be of the tolerant kind because the high virus concentration found does not necessarily relate to a high level symptoms. In the lines BPX320E 3902-01, BPX320E 3905 and BPX320F 7902 (both with Santa Clara background) only few plants showed symptoms, which can be explained by incomplete penetrance of Sw-5 gene. In the case of non-symptomatic plants, the diagnosis using DAS-ELISA revealed negative results. That outcome indicates that in some materials, where resistance is obtained from "L. peruvianum", the virus multiplication in the tissues seemed not be present.

Application of Trichoderma hamatum as a biocontroller against tomato wilt disease caused by Fusarium oxysparum F. lycopersici

The local isolate of Trichoderma hamatum had an antagonistic potential against Fusarium oxysporum f. lycopersici, the cause of tomato wilt disease. A maximum antagonism was obtained on Richard's medium at 30C and pH 6.0 in the presence of sucrose and ammonium nitrate as carbon and nitrogen sources, respectively. Scanning electron microscopy revealed that the mycoparasite [biocide, T. hamatum] grew directly towards the hyphae of the phytopathogen, followed by the formation of a contact between their hyphae, followed by the disintegration of F. oxysporum f. Lycopersici hyphae in the region of contact. T. hamatum had an extracellular chitinolytic activity in the presence of the phytopathogen as evaluated by agar plate assay. The fungicide Bavistin, the herbicide Ronstar and the insecticide malathion had an inhibitory influence on the radial growth of the phytopathogen and the antagonistic potential of T. Hamatum against the phytopathogen